Substance P exacerbates dopaminergic neurodegeneration through neurokinin-1 receptor-independent activation of microglial NADPH oxidase Chronic neuroinflammation mediated by microglia, the resident immune cells of the brain, has been implicated as a key event driving the delayed and progressive loss of dopaminergic neurons in idiopathic Parkinsons disease (PD). Post-mortem analyses of PD patients have revealed the activation of microglia and an accumulation of proinflammatory factors in the substantia nigra (SN). Similarly, Gram-negative bacterial endotoxin lipopolysaccharide (LPS) induces microglia-mediated neuroinflammation in the brains of laboratory animals and recapitulates the delayed and progressive nature of nigral dopaminergic degeneration. Previous reports have shown that humans exposed to LPS also develop Parkinsonian syndromes. However, how acute microglial activation becomes chronic and toxic to dopaminergic neurons is poorly understood. Recent findings have indicated that neurogenic signals can modulate microglial activity and function. Substance P (SP), produced in striatonigral-projecting neurons, has been shown to amplify proinflammatory responses in the CNS. This proinflammatory peptide has been implicated in the pathogenesis of PD due to its ability to stimulate dopamine (DA) release and modulate motor behaviors. However, the functional role of SP in the regulation of neuroinflammation and dopaminergic neuron survival remains elusive. Dopaminergic neurons in the SN are more susceptible to oxidative and inflammatory insults than other types of neurons. We and others have previously reported that SP is capable of stimulating or augmenting microglial activation to produce superoxide and proinflammatory cytokines in vitro. Given that SP is most concentrated in the SN, which contains a disproportionately high density of microglia (approximately 4.5 times that of other regions, we hypothesized that SP may regulate microglial activity in vivo, thereby exacerbating dopaminergic neurodegeneration in neuroinflammatory conditions. To test our hypothesis, we treated mice or various primary cell cultures with two commonly used toxins, LPS- or 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP, in animal)/1-methyl-4-phenylpyridinium (MPP+, in cell culture), which cause dopaminergic neurodegeneration. Several strains of mice deficient in SP (TAC1-/-), the SP receptor (NK1R-/-) or NADPH oxidase (NOX2; gp91phox-/-, the functional catalytic subunit of NOX2) were used. We found that SP potentiated LPS and MPTP-induced toxic effects on nigrostriatal dopaminergic neurons by enhancing microglia-mediated neuroinflammation. Importantly, we discovered that the activation of microglial NOX2 by ultra-low concentrations of SP was a novel, NK1R-independent signaling pathway responsible for the synergistic neurotoxicity induced by SP with either LPS or MPTP. These findings strongly suggest that SP facilitates neuroinflammation induced by toxins and plays a key role in damaging dopaminergic neurons via a novel NK1R-independent mechanism.